1. The Field of the Invention
The present invention relates to short interfering RNA (siRNA) that operate within the RNA interference (RNAi) pathway so as to enhance the production of industrially important polypeptides, especially biotherapeutic polypeptides and/or proteins. More particularly, synthetic siRNA that include a delivery facilitating moiety for cellular delivery and uptake can be used to enhance the production of biotherapeutic polypeptides by selectively silencing genes, a family of genes, or a biological pathway that have adverse effects on the production of the biotherapeutic polypeptides.
2. The Related Technology
Biotechnological innovations have resulted in a large number of therapeutic polypeptides (e.g., biotherapeutic agents) that are produced by recombinant nucleic acids being expressed within a cell. A primary goal of industrial producers of biotherapeutic agents is to obtain maximum yields of the polypeptide. As such, it is desirous to have cells which efficiently produce polypeptides that can be obtained in substantially pure, active, and homogeneous compositions and in quantities that are suitable for use as a biotherapeutic agent. Industrial producers of biotherapeutic agents continue to seek process improvements that allow the biotherapeutic agent to be obtained as an economically-feasible polypeptide product from the least costly configuration of cells, media, and equipment. Thus, various procedures have commonly been employed in order to optimize the production of biotherapeutic agents.
One of the procedures that has been used for increasing the efficiency of producing biotherapeutic agents is through biochemical optimization. Biochemical optimization can be considered any biochemical modulation that affects the biochemistry of the cells and/or cell cultures that are used to produce the biotherapeutic agents. Traditional biochemical optimization of cell culture bioprocesses has been used to produce therapeutic polypeptides (e.g., antibodies and other protein therapeutic agents) by modulating, and thereby optimizing the following: cell selection; cell culture conditions; cell growth media; cell culture duration; and cell growth media supplementation. These basic levers of optimization rely on fundamental nutrient biochemistry.
Another procedure that has been used for increasing the efficiency of producing biotherapeutic agents is through genetic optimization. Genetic optimization can be considered any genetic modulation that affects the genes and gene processing mechanisms of the host cells that produce the biotherapeutic agents. As such, genetically targeted modes of optimization have used the tools of molecular cloning to introduce targeted genetic alterations into the cell in order to increase expression of the desired product or to decrease the expression of genes that in any way could impair the yield of the desired biotherapeutic agent. Accordingly, genetic optimization can be employed by inserting specific sequences of DNA in the host cells or excising specific sequences of DNA therefrom in order to improve the production of the biotherapeutic agent. Such genetic optimization can be conducted by the following: regulated over-expression of apoptotic inhibitor proteins in order to decrease cell death (Van De Goor, J.; Improvement of Industrial Cell Culture Processes by Caspase-9 Dominant Negative and Other Apoptotic Inhibitors (2005) Cell Engineering Volume 4 Springer); DNA-based disruption of genes (“Knock-out”) that are found or predicted to impair efficient expression for any of a variety of reasons (Pharkya P, Maranas C D; An optimization framework for identifying reaction activation/inhibition or elimination candidates for overproduction in microbial systems. Metab Eng. 2006 January; 8(1):1-13): co-expression (“Knock-in”) of proteins that will increase expression of the desired protein, such as by agonizing or antagonizing regulatory pathways (Whitford, W.; Fed-Batch Mammalian Cell culture in Bioproduction. Bioprocess International April 2006: 31-40); and fusion of various affinity tags to the desired protein for the purpose of enhancing the speed and convenience of product purification (Hatti-Kaul, R., and Mattiasson, B.; (2003) Isolation and Purification of Proteins: Chapter 4, Genetic Approaches to Protein Purification. Marcel Dekker). Such affinity recognition elements include sequences from: immunoglobulin binding domains; repeating histidine residues; maltose binding protein, glutathione-S-transferase, and others.
While various optimization procedures have been employed in the bioproduction of biotherapeutic agents, industrial producers of biotherapeutic agents continue to seek still more optimization.